Joy N. Bailey

Relative effects of estrogen, age, and visceral fat on pulsatile growth hormone secretion in healthy women

Growth hormone (GH) secretion is subject to complex regulation. How pre- and postmenopausal age (PRE, POST), estradiol (E(2)) availability, and abdominal visceral fat (AVF) jointly affect peptidyl-secretagogue drive of GH secretion is not known. To this end, healthy PRE (n = 20) and POST (n = 22) women underwent a low- vs. high-E(2) clamp before receiving a continuous intravenous infusion of GH-releasing hormone (GHRH) or GH-releasing peptide (GHRP-2). According to analysis of covariance, PRE and POST women achieved age-independent hypo- and euestrogenemia under respective low- and high-E(2) clamps. All four of age (P < 0.001), E(2) status (P = 0.006), secretagogue type (P < 0.001), and an age x peptide interaction (P = 0.014) controlled pulsatile GH secretion. Independently of E(2) status, POST women had lower GH responses to both GHRH (P = 0.028) and GHRP-2 (P < 0.001) than PRE women. Independently of age, GHRP-2 was more stimulatory than GHRH during low E(2) (P = 0.011) and high E(2) (P < 0.001). Stepwise forward-selection multivariate analysis revealed that computerized tomographic estimates of AVF explained 22% of the variability in GHRH action (P = 0.002), whereas age and E(2) together explained 60% of the variability in GHRP-2 drive (P < 0.001). These data establish that age, estrogen status, and AVF are triple covariates of continuous peptide-secretagogue drive of pulsatile GH secretion in women. Each factor must be controlled for to allow valid comparisons of GH-axis activity.

Pre- Versus Postmenopausal Age, Estradiol, and Peptide-Secretagogue Type Determine Pulsatile Growth Hormone Secretion in Healthy Women: Studies Using Submaximal Agonist Drive and an Estrogen Clamp

CONTEXT GH-releasing peptide (GHRP), GHRH, and somatostatin are physiological regulators of pulsatile GH secretion. HYPOTHESIS Age, independently of abdominal visceral fat (AVF) and basal (nonpulsatile) GH secretion, damps pulsatile GH secretion driven by physiological (rather than pharmacological) amounts of GHRP and GHRH in an experimentally controlled estradiol (E(2)) milieu. DESIGN AND SETTING A prospectively randomized, double-blind parallel-cohort study was conducted at an academic medical center. PARTICIPANTS Community-dwelling healthy premenopausal (PRE, age 24 +/- 0.8 yr, n = 20) and postmenopausal (POST, age 63 +/- 1.8 yr, n = 22) women participated in the study. INTERVENTIONS Gonadal-axis down-regulation with leuprolide was followed by randomized addback of placebo or transdermal E(2) and separate-day iv bolus injections of a half-maximally stimulatory dose of GHRP-2 or GHRH (each 0.33 mug/kg). ANALYSIS Three-way analysis of covariance included main factors age, E(2) status, and secretagogue type and covariates AVF and basal GH secretion. RESULTS Submaximally stimulated pulsatile GH secretion was positively determined by PRE vs. POST age (P < 0.001), E(2) repletion vs. depletion (P = 0.001) and GHRP-2 vs. GHRH stimulation (P < 0.001), after adjustment for AVF and basal secretion. E(2) vs. placebo elevated fasting mean GH concentrations in both PRE and POST women (P = 0.006) but increased basal (nonpulsatile) GH secretion in PRE only (P = 0.002). PRE vs. POST age prolonged GHRH-driven GH secretory bursts by 36% (P = 0.006). CONCLUSION PRE vs. POST age, E(2) availability, and physiological peptide drive are triple determinants of pulsatile GH secretion independently of abdominal visceral fat and nonpulsatile GH secretion in healthy women.

A Pegylated Growth Hormone Receptor Antagonist, Pegvisomant, Does Not Enter the Brain in Humans

BACKGROUND GH receptors exist in the hippocampus, cerebral cortex, and hypothalamus, possibly influencing mood, cortical blood flow, and neuronal growth and mediating negative feedback. RATIONALE Pegvisomant is a recombinant mutated GH molecule with high affinity, but little or no activating capability, for the GH receptor. It is used clinically as a GH antagonist. HYPOTHESIS Systemic pegvisomant enters brain interstitial fluid via putative choroid-plexus GH receptors, thereby allowing for antagonism of central actions of GH. SUBJECTS AND LOCATION: Six adults requiring a cerebrospinal fluid (CSF) examination for nonneoplastic and noninflammatory syndromes participated at a tertiary medical center. METHODS Direct assays were conducted of serum and CSF pegvisomant concentrations 18-24 h after sc injection of pegvisomant (20 mg). OUTCOMES Median (range) concentrations of pegvisomant in serum were 215 (74-539) microg/liter and in CSF 0.035 (0.010-0.28) microg/liter (P=0.016). CSF drug levels were indistinguishable from assay threshold. Corresponding GH values were 0.29 (0.010-1.3) in serum and 0.075 microg/liter (0.01-0.13) in CSF. The geometric mean ratios of serum/CSF pegvisomant and GH concentrations were 5116:1 and 3.5:1, respectively, thus defining a more than 1400-fold difference between mutated and natural GH. CONCLUSIONS Based upon CSF measurements, a pegylated GH-receptor antagonist does not cross the human blood-brain barrier, thereby sparing inhibition of central nervous system GH actions. Thus, the capability of this antagonist to stimulate GH secretion predominantly reflects its actions outside the blood-brain barrier, such as via the median eminence and/or via suppression of IGF-I concentrations.

Estradiol supplementation in postmenopausal women attenuates suppression of pulsatile growth hormone secretion by recombinant human insulin-like growth factor type I.

BACKGROUND Why pulsatile GH secretion declines in estrogen-deficient postmenopausal individuals remains unknown. One possibility is that estrogen not only enhances stimulation by secretagogues but also attenuates negative feedback by systemic IGF-I. SITE: The study took place at an academic medical center. SUBJECTS Subjects were healthy postmenopausal women (n=25). METHODS The study included randomized assignment to estradiol (n=13) or placebo (n=12) administration for 16 d and randomly ordered administration of 0, 1.0, 1.5, and 2.0 mg/m2 recombinant human IGF-I sc on separate days fasting. ANALYSIS Deconvolution analysis of pulsatile and basal GH secretion and approximate entropy (pattern-regularity) analysis were done to quantify feedback effects of IGF-I. OUTCOMES Recombinant human IGF-I injections increased mean and peak serum IGF-I concentrations dose dependently (P<0.001) and suppressed mean GH concentrations (P<0.001), pulsatile GH secretion (P=0.001), and approximate entropy (P<0.001). Decreased GH secretion was due to reduced secretory-burst mass (P=0.005) and frequency (P<0.001) but not basal GH release (P=0.52). Estradiol supplementation lowered endogenous, but did not alter infused, IGF-I concentrations while elevating mean GH concentrations (P=0.012) and stimulating pulsatile (P=0.008) and basal (P<0.001) GH secretion. Estrogen attenuated IGF-Is inhibition of pulsatile GH secretion (P=0.042) but was unable to restore physiological GH pulse frequency or normalize approximate entropy. CONCLUSION Short-term estrogen replacement in postmenopausal women selectively mutes IGF-I-mediated feedback on pulsatile GH secretion. Disinhibition of negative feedback thus confers a novel mechanism by which estrogen may obviate hyposomatotropism.

Testosterone supplementation in older men restrains insulin-like growth factor's dose-dependent feedback inhibition of pulsatile growth hormone secretion.

BACKGROUND Pulsatile GH secretion declines in older men. The causal mechanisms are unknown. Candidates include deficient feedforward (stimulation) by endogenous secretagogues and excessive feedback (inhibition) by GH or IGF-I due to age and/or relative hypoandrogenemia. HYPOTHESIS Testosterone (T) supplementation in healthy older men will restrain negative feedback by systemic concentrations of IGF-I. SUBJECTS Twenty-four healthy men (ages, 50 to 75 yr; body mass index, 24 to 30 kg/m(2)) participated in the study. METHODS We performed a prospectively randomized, double-blind, placebo-controlled assessment of the impact of pharmacological T supplementation on GH responses to randomly ordered separate-day injections of recombinant human IGF-I doses of 0, 1.0, 1.5, and 2.0 mg/m(2). ANALYSIS Deconvolution and approximate entropy analyses of pulsatile, basal, and entropic (pattern-sensitive) modes of GH secretion were conducted. RESULTS Recombinant human IGF-I injections 1) elevated mean and peak serum IGF-I concentrations dose-dependently (both P < 0.001); 2) suppressed pulsatile GH secretion (P = 0.003), burst mass (P = 0.025), burst number (P = 0.005), interpulse variability (P = 0.032), and basal GH secretion (P = 0.009); and 3) increased secretory pattern regularity (P = 0.020). T administration did not alter experimentally controlled IGF-I concentrations, but it elevated mean GH concentrations (P = 0.015) and stimulated pulsatile GH secretion (frequency P = 0.037, mass per burst P = 0.038). Compared with placebo, T attenuated exogenous IGF-Is inhibition of GH secretory-burst mass (P < 0.038) without restoring pulse number, basal secretion, or pattern regularity. CONCLUSION The capability of systemic T to mute IGF-I feedback on pulsatile GH secretion suggests a novel mechanism for augmenting GH production.

Novel Relationships of Age, Visceral Adiposity, Insulin-Like Growth Factor (IGF)-I and IGF Binding Protein Concentrations to Growth Hormone (GH) Releasing-Hormone and GH Releasing-Peptide Efficacies in Men during Experimental Hypogonadal Clamp

BACKGROUND Sex steroids influence GH secretion in complex ways. HYPOTHESIS Analyses in a low sex-steroid milieu will help unveil the effects of age and other nonsteroidal regulators on GH secretion. CONTEXT The study was conducted in a tertiary medical center. SUBJECTS The study group included 13 healthy young men and 12 healthy older men. METHODS We used GnRH agonist-induced down-regulation of testosterone and estradiol secretion, followed by consecutive infusion of l-arginine and GHRH or GHRP-2, to test secretagogue efficacies. OUTCOMES We measured basal and pulsatile GH secretion. RESULTS During experimental testosterone/estradiol deprivation, older (57 +/- 1.7 yr) men maintained: 1) 6.8-fold less pulsatile GH secretion (P < 0.001); and 2) 2-fold lower maximal GH responses to GHRH (P = 0.0065) and GHRP-2 (P = 0.022) than young (23 +/- 1.1 yr old) individuals. Stepwise forward-selection regression analyses identified: 1) abdominal visceral fat as a dominant negative predictor of both GHRH (R(2) = 0.49; P = 0.001) and GHRP-2 (R(2) = 0.38; P = 0.005) efficacies; and 2) fasting IGF-I concentration as a major positive correlate of GHRH (R(2) = 0.52; P < 0.001) and GHRP-2 (R(2) = 0.31; P = 0.018) efficacies. Unstimulated pulsatile GH secretion was jointly correlated with IGF-I and IGFBP-3 (P = 0.039). CONCLUSION Measures of body composition (abdominal visceral fat) and pulsatile GH action (IGF-I) explain up to one half of interindividual variability in the efficacies of GHRH and GHRP-2 in sex steroid-depleted men. Accordingly, normative ranges for maximal single peptide-stimulated GH secretion in short-term hypogonadal states should incorporate the influence of these determinants as well as age.

Testosterone’s Short-Term Positive Effect on Luteinizing-Hormone Secretory-Burst Mass and Its Negative Effect on Secretory-Burst Frequency Are Attenuated in Middle-Aged Men

BACKGROUND Testosterone (T) production declines and LH pulses become smaller and more frequent in middle-aged men. The mechanisms underlying these changes are not known. RATIONALE Small frequent LH pulses in middle-aged men could reflect impaired feedback by systemic T. HYPOTHESIS Middle age disrupts negative feedback by T on selected facets of LH secretion. SUBJECTS AND SETTING Healthy men were studied at an academic medical center. METHODS The protocol comprised blockade of gonadal steroidogenesis and graded transdermal addback of T doses of 0, 2.5, 5, or 7.5 mg/d designed to span the castrate to physiological range of T concentrations in each of 23 healthy men ages 19-71 yr (interquartile range, 28-53 yr). We quantified 12-h basal and pulsatile LH secretion (92 time series) using a mathematically justified deconvolution method. RESULTS Stepwise T supplementation from the hypogonadal through the eugonadal range repressed mean (12-h) LH concentrations (P = 0.001). By regression analysis, age attenuated the capabilities of increasing T concentrations to 1) increase LH secretory-burst mass (P < 0.0001); and 2) decrease LH secretory-burst frequency (P = 0.025). Age did not alter Ts feedback on basal LH secretion, interpulse regularity, the waveform of LH secretory bursts, or the slow half-life of LH. CONCLUSION Middle age impairs both the positive and negative actions of systemic T on pulsatile LH secretion in healthy men, thus potentially explaining earlier inconsistencies in feedback studies based upon single-sample mean LH concentrations. Longitudinal studies will be required to elucidate the precise age dependence of inferred dual feedback failure.

Preservation of GHRH and GH-releasing peptide-2 efficacy in young men with experimentally induced hypogonadism.

BACKGROUND Somatostatin (SS), GHRH, GH-releasing peptide (GHRP), and the sex-steroid milieu regulate GH secretion. OBJECTIVE To test whether GHRH and GHRP remain effective secretagogs in the face of short-term hypogonadism. DESIGN Prospective, randomized double-blind. METHODS Healthy young men (n=24) received a GnRH agonist twice 3 weeks apart followed by placebo (n=13, Pl) or testosterone (n=11, testosterone) addback. SUBJECTS were then given consecutive i.v. infusions of l-arginine (to restrain SS outflow) and a maximally effective dose of GHRH or GHRP-2 (to test corresponding secretagog pathways). RESULTS GH secretion stimulated by l-arginine/GHRH and by l-arginine/GHRP-2 was unaffected by combined testosterone/estradiol (E(2)) depletion. The low testosterone/E(2) milieu decreased basal (nonpulsatile) GH secretion (P=0.038), without altering fasting pulsatile GH secretion or IGF1 or IGF-binding protein (IGFBP)-3 concentrations. IGFBP-1 (P<0.0001) and abdominal visceral fat (AVF, P=0.017) correlated negatively with fasting basal GH secretion. By contrast, IGF1 (P=0.0012) and IGFBP-3 (P=0.015) correlated positively with fasting pulsatile GH secretion. AVF (P=0.0024) was a negative determinant, and IGF1 a positive determinant (P=0.018), of GHRH-driven GH pulses. Responses to GHRP-2 were unrelated to any of these factors. CONCLUSION l-arginine/GHRP-2 appears to be an especially robust stimulus of GH secretion, since efficacy is unmodified by profound short-term hypogonadism, a range of AVF estimates, and a spectrum of IGF1, IGFBP-1, and IGFBP-3 concentrations. Whether robustness also applies to chronic hypogonadism is not known.

Regulation of basal, pulsatile, and entropic (patterned) modes of GH secretion in a putatively low-somatostatin milieu in women

Somatostatin (SS) released by hypothalamic neurons inhibits GH exocytosis noncompetitively. Therefore, we postulated that attenuation of GH feedback-induced SS outflow would help to unmask covariates of endogenous secretagogue drive. To this end, 42 healthy pre- and postmenopausal women were randomly assigned to receive leuprolide plus estradiol (E(2)) or leuprolide plus placebo. A putatively low-SS milieu was imposed by L-arginine infusion. Deconvolution and regularity analyses were applied to 6-h GH concentration-time profiles. By two-way ANOVA, age negatively (P < 0.001) and E(2) positively (P = 0.001) determined pulsatile GH secretion in the presumptively SS-deficient milieu (P < 0.001). Comparable effects were exerted on the mass of GH secreted per burst per unit distribution volume (age P = 0.001, E(2) P < 0.001, overall P < 0.001). E(2) alone predicted basal (nonpulsatile) GH secretion (P = 0.004). Stepwise forward-selection multivariate regression demonstrated that age (P = 0.0017) and E(2) (P = 0.0002) together explained 46% of intersubject variability in pulsatile GH secretion (P < 0.001) and fully replaced the negative univariate effect of abdominal visceral fat (r(2) = 0.32, P < 0.001). Moreover, age and E(2) (but not AVF) interacted to supervise GH regularity (P = 0.007). We conclude that age and E(2) availability individually and together constitute primary predictors of basal, pulsatile, and patterned GH secretion in an inferentially feedback-silenced context in healthy women. Therefore, both factors must be considered in framing hypotheses of endogenous GH drive.